Predictive Factors of Late Venous Aortocoronary Graft Failure: Ultrastructural Studies

Background

Venous aortocoronary graft arterialization may precede a preterm occlusion in some coronary artery bypass grafting (CABG) patients. The aim of the present study was to identify ultrastructural variations in the saphenous vein wall that may have an impact on the development of venous graft disease in CABG patients.

Methods

The study involved 365 consecutive patients with a mean age of 62.9±9.4 years who underwent isolated CABG. The thickness and area of the whole venous wall, the tunica intima, the tunica media and the adventitia and the number and shape (length, thickness and length/thickness ratio) of the nuclei in the medial smooth muscle cells nuclei in the distal saphenous vein segments were evaluated by ultrastructural studies. Patients were followed up for 41 to 50 months (mean 45.1±5.1). Saphenous vein graft patency was assessed by follow-up coronary angiography. Logistic regression models were used to identify independent risk factors for late graft failure.

Results

In 71 patients significant lesions in the saphenous vein grafts were observed. The whole venous wall thickness (437.5 µm vs. 405.5 µm), tunica media thickness (257.2 µm vs. 211.5 µm), whole venous wall area (2.23 mm² vs. 2.02 mm²) and tunica media area (1.09 mm² vs. 0.93 mm²) were significantly larger for this group of patients than for those without graft disease. In the latter group more elongated smooth muscle cell nuclei (higher length/thickness ratio) were found in the tunica media of the saphenous vein segments. Thickening of the saphenous vein tunica media and chunky smooth muscle cell nuclei were identified as independent risk factors for graft disease development.

Conclusions

Saphenous vein tunica media hypertrophy (resulting in wall thickening) and chunky smooth muscle cell nuclei might predict the development of venous graft disease.     

Effect of ulcerative colitis treatment on transforming growth factor beta(1) in plasma and rectal mucosa

The aim of this study was to evaluate the effect of active ulcerative colitis (UC) treatment on transforming growth factor beta(1) (TGF-beta(1)) concentration in plasma and rectal mucosa measured in 28 patients. The highest plasma values were observed in patients with the severe course of the disease (74.2+/-14.0 ng/ml), and they were significantly higher than in the group with mild one (43.7+/-5.6 ng/ml). Mean TGF-beta(1) measured in mucosal samples from patients with severe UC (563+/-146 pg/mg protein) doubled values from patients with mild UC (286+/-65 pg/mg protein). Plasma and mucosal TGF-beta(1) correlated significantly with disease activity index (DAI) and clinical activity index (CAI). Plasma TGF-beta(1) correlated additionally with scored endoscopic degree of mucosal injury. Treatment caused significant decrease of plasma and mucosal TGF-beta(1) concentrations. Patients who responded completely had higher baseline plasma and mucosal TGF-beta(1) that decreased significantly after the treatment. These results show that plasma and mucosal concentrations of transforming growth factor beta(1) are strongly associated with ulcerative colitis activity, and successful treatment of the disease results with decrease of their levels. More effective response to the treatment can be achieved in patients with higher baseline concentrations of TGF-beta(1).     

Inhibitors of bacterial N-succinyl-l,l-diaminopimelic acid desuccinylase (DapE) and demonstration of in vitro antimicrobial activity

The dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase (DapE) is a critical bacterial enzyme for the construction of the bacterial cell wall. A screen biased toward compounds containing zinc-binding groups (ZBG’s) including thiols, carboxylic acids, boronic acids, phosphonates and hydroxamates has delivered a number of micromolar inhibitors of DapE from Haemophilus influenzae, including the low micromolar inhibitor l-captopril (IC₅₀ = 3.3 μM, K_i = 1.8 μM). In vitro antimicrobial activity was demonstrated for l-captopril against Escherichia coli.     

Basic energetic parameters of Acanthamoeba castellanii mitochondria and their resistance to oxidative stress

The purpose of this study was establishing the basic energetic parameters of amoeba Acanthamoeba castellanii mitochondria respiring with malate and their response to oxidative stress caused by hydrogen peroxide in the presence of Fe(2+) ions. It appeared that, contrary to a previous report (Trocha LK, Stobienia O (2007) Acta Biochim Polon 54: 797), H(2)O(2)-treated mitochondria of A. castellanii did not display any substantial impairment. No marked changes in cytochrome pathway activity were found, as in the presence of an inhibitor of alternative oxidase no effects were observed on the rates of uncoupled and phosphorylating respiration and on coupling parameters. Only in the absence of the alternative oxidase inhibitor, non-phosphorylating respiration progressively decreased with increasing concentration of H(2)O(2), while the coupling parameters (respiratory control ratio and ADP/O ratio) slightly improved, which may indicate some inactivation of the alternative oxidase. Moreover, our results show no change in membrane potential, Ca(2+) uptake and accumulation ability, mitochondrial outer membrane integrity and cytochrome c release for 0.5-25 mM H(2)O(2)-treated versus control (H(2)O(2)-untreated) mitochondria. These results indicate that short (5 min) incubation of A. castellanii mitochondria with H(2)O(2) in the presence of Fe(2+) does not damage their basic energetics.     

Copper(II) complexes of Neobelliera Bullata Trypsin Modulating Oostatic Factor and its analogues

The stoichiometry, stability constants and solution structure of the complexes formed in the reaction of copper(II) with hexapeptide NPTNLH, i.e. the Neobelliera Bullata Trypsin Modulating Oostatic Factor (Neb-TMOF), and its analogues DPTNLH, Ac-NPTNLH and Ac-DPTNLH have been determined by potentiometric, UV-visible, CD and EPR spectroscopic methods. Upon raising pH for Ac-NPTNLH and Ac-DPTNLH peptides, copper(II) coordination starts from the imidazole nitrogen of the His⁶; afterwards three deprotonated amide nitrogens are progressively involved in metal ions coordination. In a wide pH range of 4.5-8.5 for the NPTNLH and DPTNLH ligands the CuL complex dominates with the imidazole nitrogen of His⁶ coordinated to form a macrochelate. The N-terminal amino group of the NPTNLH and DPTNLH peptides takes part in the coordination of the metal ion in the CuL, CuH₋₁L and CuH₋₂L complexes. However, at pH above 9 the CuH₋₃L complex with the {N_Im, 3N⁻} coordination mode is formed. For the CuH₋₂L complex the spectroscopic data clearly indicate the 4N {NH₂, CO or COO⁻, 2N⁻, N_Im} bonding mode with the axial coordination of the N-terminal amine group to the metal ion.     

Triterpenoids. Part 21: Oleanolic acid azaderivatives as percutaneous transport promoters

Some new oleanolic acid derivatives with lactame and thiolactame structures in the A- or C-ring were prepared and tested as percutaneous transport promoters in vitro. Their activity was comparable with activity of N-dodecylcaprolactame (Azone). A-Thiolactame derivative of methyl oleanolate (13) was the most effective compound.     

Farnesyl diphosphate synthase activity affects ergosterol level and proliferation of yeast Saccharomyces cerevisae

The yeast farnesyl diphosphate synthase (FPPS) gene was engineered so as to construct allelic forms giving various activities of the enzyme. One of the substitutions was F96W in the chain length determination region. The other, K197, conserved within a consensus sequence found in the majority of FPP and GGPP synthases, was substituted by R, E and V. An intricate correlation has been found between the FPPS activity, the amount of ergosterol synthesized and cell growth of a mutant strain defective in FPPS. About 40% of wt FPPS activity was sufficient to support normal growth of the mutant. With further decline of FPPS activity (20 down to 3%) the amount of ergosterol remained unchanged at approximately 0.16% (vs dry weight), whereas growth yield decreased and lag times increased. We postulate that, in addition to ergosterol initiating and maintaining growth of yeast cells, FPP and/or its derivatives participate in these processes.     

Hemozoin increases IFN-gamma-inducible macrophage nitric oxide generation through extracellular signal-regulated kinase- and NF-kappa B-dependent pathways

NO overproduction has been suggested to contribute to the immunopathology related to malaria infection. Even though a role for some parasite molecules (e.g., GPI) in NO induction has been proposed, the direct contribution of hemozoin (HZ), another parasite metabolite, remains to be established. Therefore, we were interested to determine whether Plasmodium falciparum (Pf) HZ and synthetic HZ, beta-hematin, alone or in combination with IFN-gamma, were able to induce macrophage (Mphi) NO synthesis. We observed that neither Pf HZ nor synthetic HZ led to NO generation in B10R murine Mphi; however, they significantly increased IFN-gamma-mediated inducible NO synthase (iNOS) mRNA and protein expression, and NO production. Next, by investigating the transductional mechanisms involved in this cellular regulation, we established that HZ induces extracellular signal-regulated kinase (ERK)1/2 mitogen-activated protein kinase phosphorylation as well as NF-kappaB binding to the iNOS promoter, and enhances the IFN-gamma-dependent activation of both second messengers. Of interest, cell pretreatment with specific inhibitors against either NF-kappaB or the ERK1/2 pathway blocked the HZ + IFN-gamma-inducible NF-kappaB activity and significantly reduced the HZ-dependent increase on IFN-gamma-mediated iNOS and NO induction. Even though selective inhibition of the Janus kinase 2/STAT1alpha pathway suppressed NO synthesis in response to HZ + IFN-gamma, HZ alone did not activate this signaling pathway and did not have an up-regulating effect on the IFN-gamma-induced Janus kinase 2/STAT1alpha phosphorylation and STAT1alpha binding to the iNOS promoter. In conclusion, our results suggest that HZ exerts a potent synergistic effect on the IFN-gamma-inducible NO generation in Mphi via ERK- and NF-kappaB-dependent pathways.     

The influence of intracellular idarubicin and daunorubicin levels on drug cytotoxicity in childhood acute leukemia

Uptake and efflux of two anthracyclines, idarubicin (IDA) and daunorubicin (DNR), was studied in childhood acute leukemia samples. A comparison of IDA and DNR transport phenomena in relation to drug cytotoxicity and expression of P-glycoprotein (PGP) was made. Intracellular content of IDA/DNR was determined by flow cytometry using the fluorescent properties of the drugs. In vitro drug cytotoxicity was measured by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) assay. PGP expression was analysed by flow cytometry. The uptake and efflux rates were non-significantly higher for IDA than DNR. There were no differences between three types of leukemia with respect to drug content during accumulation and retention. After correction for the cell volume, intracellular concentration of both drugs in each moment of uptake and efflux was significantly lower in relapsed ALL and AML samples in comparison with initial ALL cells. Efflux, but not uptake, of both drugs was inversely correlated with PGP expression and IDA, but not DNR, cytotoxicity. The cytotoxicity was correlated with drug accumulation for both drugs and with drug retention for IDA. In conclusion, it seems that (1) intracellular content was related to the lipophilic properties of the drugs rather than to the type of leukemia, (2) decreased intracellular concentration of both drugs might have an impact on compromised therapy results in AML and relapsed ALL children, (3) IDA presents higher cytotoxicity, which possibly might be decreased by the presence of PGP. These results might have a practical impact on the rational design of new chemotherapy protocols.     

In vitro activity of oxazaphosphorines in childhood acute leukemia: preliminary report

Glufosfamide (beta-D-glucosyl-ifosfamide mustard) is a new agent for cancer chemotherapy. Its pharmacology is similar to commonly used oxazaphosphorines, but it does not require activation by hepatic cytochrome P-450 and preclinically demonstrates lower nephrotoxicity and myelosuppression than ifosfamide. The aim of the study was a comparison of the drug resistance profiles of glufosfamide and other oxazaphosphorines in childhood acute leukemias. Leukemic cells, taken from children with ALL on diagnosis (n = 41), ALL on relapse (n = 12) and AML on diagnosis (n = 13) were analyzed by means of the MTT assay. The following drugs were tested: glufosfamide (GLU), 4-HOO-ifosfamide (IFO), 4-HOO-cyclophosphamide (CYC) and mafosfamide cyclohexylamine salt (MAF). In the group of initial ALL samples median cytotoxicity values for GLU, IFO, CYC and MAF were 15.5, 33.8, 15.7 and 7.8 microM, respectively. In comparison with initial ALL samples, the relative resistance for GLU and IFO in relapsed ALL samples was 1.9 (p = 0.049) and 1.3 (ns), and in initial AML samples 31 (p < 0.001) and 5 (p = 0.001), respectively. All oxazaphosphorines presented highly significant cross-resistance. Glufosfamide presented high activity against lymphoblasts both on diagnosis and on relapse.